Project Summary Many affective disorders are caused or exacerbated by exposure to severe or repeated stressors. Despite the important role of stressor exposure in modulating emotion, the mechanisms by which central emotional circuits are altered by stress are still unknown. Substantial evidence has suggested that the dorsal anterior bed nucleus of the stria terminalis (BNST) mediates anxiety-like behavior in humans and animals, and it is likely that altered BNST function underlies anxiety disorders. We have shown that pituitary adenylate cyclase activating polypeptide (PACAP) activation and release in the BNST mediates many of the behavioral effects of repeated stress in males and females, and that circulating PACAP levels and a unique single nucleotide polymorphism in the PAC1 receptor predict PTSD symptoms and diagnosis in women, suggesting that PACAP systems. In non-stressed organisms, BNST PACAP release likely originates from the lateral parabrachial nucleus (LPBn); however, we argue that following chronic stress, local BNST PACAP production and release is increased concurrent to an increase in PAC1 receptor-mediated activation of locally-projecting corticotropin-releasing factor (CRF)-expressing neurons in the BNST to produce pathological anxiety. Importantly, different PAC1 receptor isoforms can signal via multiple mechanisms to produce short-duration and sustained effects on neuronal excitability. Hence, the activation of cAMP subsequent to membrane-bound PAC1 receptor activation of adenylyl cyclase (AC) likely leads to the immediate changes in BNSTov excitability observed following PACAP. However, the phosphorylation and activation of extracellular signal-regulated kinase 1/2 (pERK) via endosomal signaling likely leads to a sustained anxiogenic response, and pERK signaling may also support trophic actions to enhance anxiety-like behavioral responding for even longer periods. The experiments in this application use a combination of molecular, physiological and behavioral approaches to investigate whether PACAP targets different populations of BNSTov neurons in stress- and unstressed males and females, and whether different PAC1 receptor signaling pathways mediate anxiogenic behavior with different temporal characteristics. Understanding the signaling cascades and cellular targets that mediate the effects of BNST PACAP could help to better target the maladaptive consequences of stressor exposure.